The objective of the Program is to study the molecular regulation of pattern formation in the developing vertebrate limb, and to begin to apply this basic knowledge to understanding the nature of human genetic disorders affecting limb patterning. In Project 1, the roles and relationships between the homeobox-containing genes Msx2 and D1x-5 in the regulation of activity of the apical ectodermal ridge (AER), which plays a crucial role in directing outgrowth and patterning of the lib, will be studied, as will their roles in specification of the anterior mesoderm of the limb bud. The role of D1x-5 in regulating the onset of limb cartilage differentiation and cartilage-specific gene expression will also be investigated. In Project 2, the roles of the growth factors IGF-I and insulin in the reciprocal interactions between the AER and subridge mesoderm that are responsible for outgrowth of the limb will be studied, as will the roles of these growth factors in the initial formation of the limb bud from the somatopleure of the lateral plate of the chick embryo. In Project 3, an in-depth analysis of the transcriptional mechanisms regulating the tissue-specific expression of msx2 in the AER and anterior mesoderm of the developing limb will be undertaken using reporter constructs in transgenic mice to provide insight into how the msx2 gene is regulated and how the regulatory network is involved in pattern formation. In Project 4, genetic linkage analyses will be used to map the loci of the human chondrodysplasias, Grebe syndrome and ectrodactyly, which are human genetic disorders in which the clinical manifestations suggest errors in developmental patterning of the limbs. A Core unit will supply Projects 1-3 with amelic and polydactylous mutant chick embryos that will be used in the analysis of the roles of regulatory genes in patterning. The integrated goal of this Program is to investigate the nature, regulation, interactions, and mechanisms of action of some of the genes and signalling molecules involved in the patterning of the chick limb, and to use this knowledge to understand human genetic disorders of the limb.